The physical-chemical properties and biological roles of lipid membranes are being investigated by atomic force microscopy (AFM) and other technologies in this project via several collaborations. (1) In a continuing work with NIAID collaborators (Drs. J.A. Dvorak and F. Tokumasu) on heterogeneities of lipid membranes, we have extended our quantitative studies of the main phase transition in 2-dimyristoyl-sn-glycero-phosphocholine (DMPC), revealing an intrinsic domain size of about 4.2 nm in diameter, to tri-lipid mixture system of dipalmitoyl phosphatidylcholine (DPPC), dilauroyl phosphatidylcholine (DLPC), and cholesterol (chol). This tri-lipid system mimics more closely biological membranes and has been previously investigated via light microscopy by Prof. G. W. Feigenson (Cornell University) who is also a collaborator for the current study. For this tri-lipid system, we quantified a moderate membrane-mica attractive support from large unilamellar vesicle (LUV) surface adsorption and fusion. Then, our AFM imaging revealed extensive nanoscopic membrane domains (c.f. membrane rafts) that range from 26 to 38 nanometers depending on cholesterol level, DLPC and DPPC ratio, and other phase space parameters. Our overall aim is to apply novel AFM imaging and mathematical analyses toward a comprehensive understanding of the cellular membrane under influence of its composition and native interactions. (2) Also with LPD, NIAID collaborators (Drs. J.A. Dvorak and T. Arie), we developed new video microscopy and image analyses to characterize the red blood cell flicker and edge dithering phenomena during the Plasmodium falciparum malaria infection process. We have found that the parasitic infection markedly modifies cell membrane dynamics and are quantifying these changes according to parasitic developmental stages, in potential relevance to malaria disease mechanism in human microcirculations. (3) In a further expansion of above two collaborations and with in addition NIAAA research collaborators (Drs. B. J. Litman and S.-L. Niu), we are using AFM and Differential Scanning Calorimetry (DSC) to characterize micro-domain and nanoscopic structures in a new three-component DDPC/DPPC/cholesterol system mimicking rod outer segment membranes in the vision pathway. Unusual membrane properties are being identified and correlated to presence of the double unsaturated acyl chains in the DDPC lipid. (4) Finally, with another NIAID collaborating scientist (Dr. J. Silver), nanoscopic energetic and structural descriptions of membranes are being developed to understand fusion pore dynamics important to virus invasion and cellular trafficking processes. The potential role of membrane lateral organizations in these processes is yet to be understood.